Molecular Determinants of Calcitic Biomineralization.

方解石生物矿化的分子决定因素。

• 批准号: RGPIN-2016-04410
• 负责人: Hincke, Maxwell
• 金额: $ 2.99万
• 依托单位: University of Ottawa
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=653601
• 关键词: Molecular; Determinants; Calcitic; Biomineralization.

Flaws in chicken eggshell fabrication are a human health and food safety issue; the nutritious table egg is an important source of protein, minerals and vitamins for 100's of millions of individuals living in poverty worldwide. In Canada it is estimated that about 10,000 annual cases of salmonella poisoning are due to contamination of cracked eggshells that find their way into the food chain. In this sense, the reproducible nature of calcitic biomineralization of the eggshell is of critical concern to human society. My research program during the past 20 years has investigated the integrated defense strategies that function at this biomineralized barrier. My studies have revealed how protein components of the eggshell matrix modulate calcium carbonate crystal nucleation and growth in order to understand regulation of eggshell mechanical properties. Moreover, we have demonstrated the dual function of antimicrobial eggshell matrix proteins (EMPs) that regulate mineralization of the shell barrier which maintains the pathogen-free status of the egg.***The goals are to investigate the initiation of eggshell mineralization and the regulatory role of proteins. The role of specific EMPs and amorphous calcium carbonate (ACC) in eggshell formation (mineralization) and resorption (demineralization) will be determined. The dual role of antimicrobial EMPs will be further investigated.****Specific objectives:****1. Our transcriptomic analysis of specific gene expression in the isthmus segment of the oviduct identified collagen X (COL10A1), fibrillin-1 (FBN1) and cysteine rich eggshell membrane protein (CREMP) as key structural constituents. Their incorporation and relative organization in the native eggshell membranes will be investigated using specific antibodies. The assembly of these proteins into stable fibres will be investigated in vitro with purified native and/or recombinant proteins.****2. My hypothesis that the mammillary cone contains a stabilized pool of ACC as a Ca reservoir that the embryo accesses during growth will be tested by investigating the status of CaCO3 and identifying associated proteins in the calcium reserve body at the mammillary cone base.****3. Candidate proteins (such as OC17, NUCB) proposed to modulate mineralization during eggshell formation will be investigated with in vitro crystallization studies. Regulatory molecules will be validated by reconstitution experiments utilizing purified recombinant proteins to modulate the ACC to calcite transformation, and to reproduce nucleation of mammillary cones.****4. The antimicrobial activity and CaCO3 nucleation / crystal growth properties of OCX32 will be investigated to assess the dual function of this important matrix protein.****These studies will provide solutions to serious food safety issues such as salmonella contamination of the table egg.**

鸡蛋壳制造中的缺陷是人类健康和食品安全问题;营养丰富的餐桌鸡蛋是世界各地生活在贫困中的数亿人的蛋白质，矿物质和维生素的重要来源。据估计，在加拿大，每年约有10，000例沙门氏菌中毒病例是由于破裂的蛋壳污染而进入食物链造成的。从这个意义上说，蛋壳的方解石生物矿化的可再生性对人类社会至关重要。在过去的20年里，我的研究项目调查了在这种生物矿化屏障上发挥作用的综合防御策略。我的研究揭示了蛋壳基质中的蛋白质成分如何调节碳酸钙晶体的成核和生长，以了解蛋壳机械性能的调节。此外，我们已经证明了抗微生物蛋壳基质蛋白（EMP）的双重功能，该蛋白调节壳屏障的矿化，从而维持鸡蛋的无病原体状态。目的是研究蛋壳矿化的启动和蛋白质的调节作用。将确定特定EMP和无定形碳酸钙（ACC）在蛋壳形成（矿化）和再吸收（脱矿）中的作用。抗菌电磁脉冲的双重作用将得到进一步研究。*具体目标：我们对输卵管峡部段特异性基因表达的转录组学分析确定了胶原X（COL10A1）、胶原蛋白-1（FBN1）和富含半胱氨酸的蛋壳膜蛋白（CREMP）为关键结构成分。将使用特异性抗体研究它们在天然蛋壳膜中的掺入和相对组织。将在体外用纯化的天然和/或重组蛋白研究这些蛋白质组装成稳定的纤维。* 2.我的假设，即乳头锥包含一个稳定的ACC池作为胚胎在生长过程中访问的钙库，将通过研究CaCO3的状态和鉴定乳头锥基部钙储备体中的相关蛋白来测试。3.候选蛋白质（如OC 17，NUCB）建议在蛋壳形成过程中调节矿化将在体外结晶研究。调节分子将通过利用纯化的重组蛋白的重建实验来验证，以调节ACC到方解石的转化，并再现乳头状视锥的成核。**** 4.将研究OCX 32的抗微生物活性和CaCO3成核/晶体生长特性，以评估这种重要基质蛋白的双重功能。*这些研究将为严重的食品安全问题提供解决方案，例如餐桌鸡蛋的盐污染。